have you ever used hygromycin to screen transfected HepG2 or gastric cancer cell - all the cells died, why? (Nov/19/2006 )

hi, all, I used hygromycin B to screen stable transfected HepG2 cells and gastric cancer cell, but after I tried a serial of concentrations , from 20ug/ml to 200ug/ml, finally all the cells were dead. I thought some cells were still alive after 25 days screening( at least they looked good), but after tripsinizing the cells , all the cells detached from the plate and looked not as good as before, I am afraid they are also going to die. what's the problem? has hygromycin have a characteristic to make cells look like this?

-bigbeauty-

you dont need to screen for 25 days. One week kill curve is more than sufficient. Why dont you go lower than 20 mcg/ml and see what happens.